Extended Data Fig. 2: Validation of the temporal information in the UMAP by using complementary tools.
From: A root phloem pole cell atlas reveals common transcriptional states in protophloem-adjacent cells
a) Color-coded UMAP according to the longitudinal sections in Brady et al. 2007, showing a developmental progression in each cell lineage. b) Bar plot indicating the percentage of cells contributed by each cluster to each of the Slingshot trajectories shown in the main text. Bars are coloured by trajectory. c) RNA velocity analysis using scVelo, with velocity vectors projected on our UMAP d) Confocal pictures of pAPL::VENUSer showing continuous expression in PSE in the early root meristem with a patchy expression in the neighbouring cells that gets stable in CC and MSE shootward. As observed in the zoom picture, the signal in PPP gets weaker after PSE enucleation. The pictures are accompanied by a UMAP showing APL cluster-weighted normalised expression in the phloem pole cell atlas. Scale bar in the longitudinal sections is 25 µm while it is 10 µm in the cross sections and zoom. White arrowheads point to PSE cells as a reference point. 25 independent seedlings from three different lines expressing this construct were observed e) Probability of a cell to be assigned to different trajectories, ranging from 0 to 1. In the image we are showing a few clusters as an example. f) The expression of APL and PSE enucleation markers NAC086 and NEN4 was plotted along the PPP, CC and PSE trajectories, with the cells coloured by cluster number in the UMAP. NAC086, an APL target, appears later than APL in the PSE trajectory, and NEN4, a NAC086 target, appears later than NAC086, indicating our approach matches the temporal aspects observed in PSE biology in the roots. The dots representing the cells are coloured according to cluster colours in panel c and Fig. 1b. The black line is a smoothed trend estimated from a non-parametric generalised additive model. g) Initiation of S17 and SAPL gene expression. Distance from QC (µm) was measured to the first cell expressing each gene in 7 days post sowing (dps) seedlings, n = 10 for S17 and n = 11 for SAPL.
